Project Summary: The goal of this project is to develop the first negative allosteric modulator (NAM)-based positron emission tomography (PET) ligand for metabotropic glutamate receptor subtype 2 (mGlu2) based on negative allosteric modulation as imaging biomarker for drug discovery. The physiological function of mGlu2 is to negatively regulate endogenous glutamate release and protect neurons against excitotoxicity. Continuous efforts to modulate mGlu2 represent a broad pharmacological approach for the treatment of CNS disorders, including drug addiction, depression, schizophrenia, Parkinson?s disease and Alzheimer?s disease. As a non-invasive imaging technology, PET is capable of quantifying biochemical processes in vivo, and a suitable mGlu2 PET ligand would substantially improve our understanding of mGlu2-based glutamatergic signaling under normal and disease conditions otherwise inaccessible by ex vivo (destructive) analysis. Furthermore, quantification of mGlu2 by PET would provide the assessment of distribution, target engagement and dose occupancy of mGlu2-targeted new neurotherapeutics. To date there is no successful examples have been demonstrated to image mGlu2, representing a significant deficiency of our ability to study this target in vivo. Therefore, we propose to develop a PET ligand for mGlu2 that will overcome the two major shortcomings of previous PET ligands (low brain uptake and limited target specificity) and be the first mGlu2 imaging agent for drug discovery and clinical translation. The first NAM-based mGlu2 ligand, [11C]QCA, was recently developed by the PI at MGH. [11C]QCA showed excellent in vitro specific binding and target selectivity towards mGlu2, but was discontinued due to low brain penetration. In our second generation mGlu2 NAMs, we identify a lead pyridine compound with high potency and selectivity towards mGlu2. A 11C-isotopolgue of this lead showed reasonable brain uptake (1.1 SUV) and target engagement in pretreatment studies (~60% blockade). Though our lead candidate is a promising lead for the development of new mGlu2-targeted target engagement tool, PET ligands with higher brain penetration, improved potency and selectivity, increased specific binding ratios are needed for optimal imaging and quantification of mGlu2. In this proposal, we will design and prepare a series of carefully chosen mGlu2 NAMs, radiolabel top candidates with 11C or 18F, and evaluate their ability to quantify mGlu2 activity and changes during drug challenges in rodents and nonhuman primates. The impact of this work is not only to develop the first potent and selective mGlu2 PET NAM ligand for drug discovery, but also ultimately, to prepare this imaging tool for potential clinical translation and monitor direct effect of mGlu2 therapeutic agents in human brain. The target validation and occupancy for efficacious mGlu2- related treatment can also be determined and quantified by the mGlu2 PET ligands in this work. Relevance: This proposal has the potential to improve public health and help patients suffering from CSN disorders through the discovery of neurotherapeutics using mGlu2 PET ligands.